This invention relates to a medical diagnostic method and, more particularly, to an in vitro diagnostic method of screening for immune deficiency in nephrotic patients.
The nephrotic syndrome (NS) is characterized by extensive urinary loss of albumin leading to hypoalbuminemia, edema and hypercholesterolemia. See, for example, Brenner and Stein, Contemporary Issues in Nephrology, Vol. 9, Nephrotic Syndrome, Churchill Livingston, New York, 1982, pp. 1-308. Patients with this syndrome frequently have suppressed clinical and in vitro immune responsiveness; the disease in which this 0henomenon has been most extensively studied is minimal change nephrotic syndrome (MCNS). Unique attributes of MCNS include the lack of morphological changes sufficient to account for proteinuria, the observed pattern of glomerular permselectivity, a frequent association with antecedent immunogenic stimuli, and the marked responsiveness of this disease to immunomodulatory agents. Patients with MCNS show evidence of decreased cellular immunity. Furthermore, their sera have been found to suppress lymphocyte proliferation and to show cytotoxic activity. Enhanced suppressor cell function has also been described. Thus, clinical evidence of altered immunity appears to be associated with increased suppressor cell activity.
Patients with nephrotic syndrome frequently have suppressed immune responsiveness by both clinical and in vitro evaluation. Thus, bacterial infection, particularly primary peritonitis, frequently occurs during relapse. MCNS patients also have decreased skin reactivity to tuberculin and dinitrochlorobenzene which improves with remission of symptoms. Elevated percentages of both T and B lymphocytes have been demonstrated by techniques of examining either rosetting or cell surface marker expression. In addition, serum immunoglobulin levels are abnormal, with relapse-associated increased IgM and decreased IgG levels being reported in MCNS and other forms of nephrotic syndrome as well. Specific titers may also be affected, since a patient has been reported to show an appropriate initial response to pneumococcal polysaccharide vaccine but a subsequent rapid decrease in titer.
More recently, immune responses in nephrotics have been evaluated using in vitro techniques. Patient sera have been found to be toxic to control lymphocytes; increased serum migration inhibitory factor activity, and increased monocyte cytotoxicity against cultured renal epithelial cells have also been described. Several studies have shown decreased lymphocyte proliferation in the presence of sera from MCNS patients in relapse. A heat-stable substance in patient serum has been described which binds to normal lymphocytes and decreases the proliferative response to mitogens. Increased Concanavalin A-activated suppressor cell activity has also been reported in MCNS patients in relapse compared to healthy controls, patients in remission, and patients with glomerulonephritis. Although these findings appear most striking in MCNS, they may not be restricted to minimal change disease; several of the studies mentioned above found similar abnormalities in other forms of nephrotic syndrome.
Despite this considerable body of information, the mechanism of immune suppressive activity is unclear. Although the basic mechanism of suppressed responses in MCNS remains unclear, suppressor cells have been shown in most basic systems to act through release of soluble suppressor factors. One such system which has been fairly well-characterized is the soluble immune response suppressor (SIRS) pathway. SIRS is a product of mitogen- or interferon-activated murine or human suppressor T lymphocytes which inhibits in vitro production of antibody when added at or near initiation of lymphocyte cultures. See, for example, Rich and Pierce, J. Immunol. 112, 1360-68 (1974); and Aune and Pierce, Proc. Natl. Acad. Sci. USA 79, 3808-12 (1982). Suppression also occurs when factor is added late in the culture period after activation to its suppressive form, SIRS.sub.ox, by reaction with low concentrations of H.sub.2 O.sub.2, as described by Aune and Pierce, Lymphokines, Vol. 9, E. Pick (ed), Academic Press Inc., New York, 1984, pp. 257-77. Human SIRS has a molecular weight of 110-150,000 daltons when fractionated by chromatography on Sephracryl.RTM.-200 gel using buffers of physiologic ionic strength and 10-15,000 daltons when eluted in high ionic strength buffers. It is acid and protease sensitive, and its in vitro activity is blocked by levamisole and catalase, which block activation of SIRS to SIRS.sub.ox, and by 2-mercaptoethanol (2-ME), which inactivates SIRS.sub.ox. Such characterization of human SIRS is disclosed by Schnaper, Pierce and Aune, J. Immunol. 132, 2429-35 (1984). Inhibition of suppression by these reagents and activation by peroxide are characteristic of both murine and human SIRS and are useful techniques in screening for the presence of SIRS in other systems. For further background information on the purification and characterization of SIRS, see also Aune, Webb and Pierce, J. Immunol. 131, 2848-52 (1983).
In view of the foregoing, it is believed that it would be useful to be able to relate immune suppression to disease etiology and that a diagnostic method of screening for immune deficiency in nephrotic patients would be of considerable importance to establish such relation.